Role of proteoglycan sulfation during muscle regeneration in dystrophic animals

蛋白多糖硫酸化在营养不良动物肌肉再生过程中的作用

基本信息

批准号：
9066087
负责人：
Matthew Tierney
金额：
$ 0.95万
依托单位：
SANFORD BURNHAM PREBYS MEDICAL DISCOVERY INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-06-01 至 2016-09-23
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9066087
关键词：
Adult Animals Behavior Behavioral Binding Biological Assay Biological Availability Cells Communication Complex Cues Development Disease Progression Donor person Duchenne muscular dystrophy Environment Event Exhibits FGF2 gene Fetal Tissues Fibroblast Growth Factor Fibroblast Growth Factor Receptors Fluorescence-Activated Cell Sorting Gene Expression Goals Growth Health Heparan Sulfate Proteoglycan Impairment In Vitro Intervention Ligands Measures Mediating Mediator of activation protein Membrane Mitogens Molecular Monitor Mus Muscle Muscle satellite cell Muscular Atrophy Natural regeneration Pathway interactions Patients Pattern Phosphorylation Play Post-Translational Protein Processing Pre-Clinical Model Proteoglycan Quality of life Regulation Resistance Role Signal Pathway Signal Transduction Signaling Molecule Skeletal Muscle Staging Stem cells Sulfatases Testing Therapeutic Effect Transplantation Wasting Syndrome Western Blotting Work beta catenin bioluminescence imaging cell behavior extracellular fetal functional disability human disease improved in vivo innovation insight mouse model muscle regeneration myogenesis neutralizing antibody new therapeutic target pre-clinical prospective receptor regenerative repaired response self-renewal sulfation syndecan therapy design tool

项目摘要

DESCRIPTION (provided by applicant): Muscle stem cells (MuSC) are required for the development and repair of skeletal muscle. During these dynamic periods of muscle growth and regeneration, MuSC must selectively interpret a host of signaling molecules to self-renew. This ability is progressively impaired in Duchenne muscular dystrophy (DMD), a devastating muscle wasting disease for which there is no cure. DMD is characterized by altered microenvironmental signaling that negatively influences MuSC-mediated regenerative potential. To better understand how MuSC communicate with their extracellular environment, I explored several membrane-bound mediators of signal transduction. Heparan sulfate proteoglycans (HSPG), in particular syndecans, are highly expressed in MuSC and can regulate ligand bioavailability and receptor formation. Heterogeneous sulfation patterns on HSPG via post-translational modifications increase behavioral complexity, leading to a differential activity of critical signalng pathways. Because regeneration in the adult recapitulates several aspects of myogenesis, I focused my preliminary studies on HSPG sulfation in fetal MuSC, the developmental precursors of adult MuSC. I have demonstrated that Sulf1, a specific regulator of HSPG 6-O-sulfation, is downregulated in fetal MuSC. I provide evidence that fetal MuSC are resistant to myogenic commitment in vitro and capable of rapid in vivo expansion and long-term self-renewal following transplantation. Furthermore, uncommitted fetal MuSC are preferentially sensitive to the potent mitogen FGF2 while less responsive to Wnt/β-catenin-mediated myogenic differentiation. Indeed, Sulf1 has been previously shown to simultaneously promote Wnt/β-catenin and inhibit FGF2 signaling. Sulf1 and HSPG expression are dysregulated in muscle wasting conditions. Therefore, the goal of this project is to examine the role of HSPG 6-O-sulfation on the regulation of dystrophic MuSC self-renewal and regenerative potential. To accomplish this, I propose (Aim 1a) to validate HSPG sulfatase and FGF and Wnt/β-catenin signaling component expression levels throughout myogenesis and in dystrophic animals. To determine MuSC responsiveness to FGF and Wnt/β-catenin pathway activation (Aim 1b), I will measure the expression and phosphorylation levels of downstream signaling pathway effectors. Inhibition of HSPG 6-O-sulfation will ascertain its role in FGF receptor complex formation and Wnt3a ligand bioavailability, defining the mechanisms underlying HSPG regulation of proper pathway activation. (Aim 1c). I will determine the impact of Sulf1 inhibition on dystrophic, myofiber-associated MuSC self-renewal (Aim 2a). Finally, I will evaluate the functional role of HSPG 6-O-sulfation in a preclinical model of DMD by inhibiting Sulf1 expression in (Aim 2b) transplanted, donor MuSC and (Aim 2c) native, dystrophic MuSC. Through these studies I will investigate a molecular mechanism, HSPG 6-O-sulfation, able to regulate FGF and Wnt/β-catenin signaling and MuSC regenerative potential in dystrophic animals. This work will potentially identify Sulf1 as a novel therapeutic target for the enhancement of MuSC self-renewal and the amelioration of DMD.

描述（由适用提供）：肌肉干细胞（MUSC）是骨骼肌发展和修复所必需的。在这些动态的肌肉生长和再生时期，MUSC必须选择性地解释大量信号分子以自我更新。这种能力逐渐受损，在肌肉营养不良（DMD）中，这是一种毁灭性的肌肉浪费疾病，无法治愈。 DMD的特征是微环境信号改变，对MUSC介导的再生潜力产生负面影响。为了更好地了解MUSC如何与其细胞外环境进行沟通，我探索了几个信号转导的膜结合的介质。硫酸乙酰肝素蛋白聚糖（HSPG），特别是Syndecans，在MUSC中高度表达，可以调节配体生物利用度和受体形成。通过翻译后修饰在HSPG上的异质硫酸化模式增加了行为复杂性，从而导致关键信号途径的不同活性。由于成年人的再生概括了肌发生的几个方面，因此我将初步研究集中在胎儿MUSC（成人MUSC的发育前体）中的HSPG硫酸化上。我已经证明，在胎儿MUSC中，Sulf1是HSPG 6-O-硫化的特定调节剂。我提供的证据表明，胎儿MUSC对体外的肌源性承诺有抵抗力，并且能够在移植后能够快速的体内扩张和长期自我更新。此外，未承诺的胎儿MUSC对潜在的有丝分裂原FGF2更敏感，而对Wnt/β-catenin介导的肌源性分化的反应较低。实际上，先前已证明Sulf1仅促进Wnt/β-catenin并抑制FGF2信号传导。在肌肉浪费条件下，SULF1和HSPG表达失调。因此，该项目的目的是检查HSPG 6-O-硫化对可营养不良的MUSC自我更新和再生潜力的作用。为此，我建议（AIM 1A）验证HSPG硫酸硫酸硫酸硫磺酶，FGF和Wnt/β-catenin信号传导成分在整个肌发生和营养不良动物中的表达水平。为了确定MUSC对FGF和Wnt/β-catenin途径激活的反应性（AIM 1B），我将测量下游信号通路效应的表达和磷酸化水平。 HSPG 6-O-硫化的抑制作用将确定其在FGF受体复合物形成和WNT3A配体生物利用度中的作用，从而定义了适当途径激活的HSPG调节基础的机制。（AIM 1C）。我将确定Sulf1抑制对营养不良的，肌纤维相关的MUSC自我更新的影响（AIM 2A）。最后，我将通过抑制（AIM 2B）移植的供体MUSC和（AIM 2C）天然肌营养不良的MUSC中的Sulf1表达来评估DMD临床前模型中HSPG 6-O-硫化的功能作用。通过这些研究，我将研究一种分子机制，即HSPG 6-O-硫化，能够调节营养不良动物中FGF和Wnt/β-catenin信号传导和MUSC再生潜力。这项工作将有可能将Sulf1识别为增强MUSC自我更新和DMD改善的新型治疗靶标。